Dephosphorylation-controlled extraction of phosphorylatable psychoactive alkaloids

ABSTRACT

This invention relates to the extraction of psychoactive compounds from fungus for use in medicine. Raw fungus is dried and ground. The solvent used for extraction is methanol or a hydro-methanol mixture, an acidic hydro-methanol mixture, or an alkaline hydro-methanol mixture. The extraction slurry is filtered and pH-adjusted if necessary. The methanol in the solvent is then completely evaporated and water added back, where necessary, to form a concentrated slurry. The concentrated slurry is then standardized to provide a known concentration of the psychoactive alkaloids that have been extracted. The standardized slurry may then be dried to result in a powdered extract with a precisely defined purity of psychoactive compounds.

RELATED APPLICATIONS

This application is a continuation of U.S. patent application Ser. No.17/841,323, filed on Jun. 15, 2022, which is a continuation ofInternational Application No. PCT/CA2021/050823 filed Jun. 16, 2021,which claims the benefit of U.S. Provisional Patent Application No.63/040,317 filed on Jun. 17, 2020, and U.S. Provisional PatentApplication No. 63/046,089 filed on Jun. 30, 2020. This application alsoclaims priority to Canadian Application No. 3089455 filed Aug. 7, 2020,Canadian Application No. 3088384 filed Jul. 29, 2020, InternationalApplication No. PCT/CA2021/050813 filed Jun. 14, 2021, InternationalApplication No. PCT/CA2021/050822 filed Jun. 16, 2021, each of which areincorporated by reference in their entireties.

TECHNICAL FIELD

This application relates to the extraction of active ingredients fromfungus. More specifically, it relates to extracting psychoactivecompounds from fungus and forming an extract of known purity.

BACKGROUND

Varieties of mushrooms have played important roles in most societies.The active ingredients in mushrooms, especially psilocybin mushroomswith psychoactive compounds such as psilocybin, psilocin, baeocystin,norbaeocystin, ibotenic acid, and norpsilocin, have been found to havemedicinal properties including relief of symptoms of various diseasesand conditions. The concentration of active psilocybin mushroomcompounds varies not only from species to species, but also frommushroom to mushroom inside a given species, subspecies or variety. Thesame holds true even for different parts of the same mushroom ormycelium.

Various methods of extraction, which have been used to separate naturalextracts from a variety of mushrooms, have resulted in difficulties withlarge crop-to-crop variability. This is as well as the problem of alarge variability within a single plant or fungus in terms of theconcentration of the active psychoactive compound and its stability.Different solvent choices extract the psychoactive compounds equally,some of them electively extract one or the other, and some convert thecompounds between each other or degrade them into non-psychoactivecompounds. Many extraction processes for extracting standardizedconcentrations of the compounds for direct medical use are usuallycomplex. This results in expensive extraction processes and a high costof isolated, natural extracts.

U.S. Pat. No. 3,183,172 to Heim et al. relates to an industrial processfor the isolation of active compounds from mushrooms grown underpredetermined conditions. With the predetermined growing conditions,mushrooms grow with ten times more active mycelium and sclerotium, andincreased concentrations of psychoactive compounds. However, a largeportion of the target compounds are lost during the extraction processor not extracted at all. This problem is significant with respect tovery potent extracts of psilocybin mushrooms, considering that a normaldose for use ranges from only 5 mg to 25 mg. The extracted psychoactivecompounds are generally without a stable and standardized concentration.

To date, the focus has largely been on synthetic preparations of thesecompounds because of the many difficulties associated with naturallyextracted preparations. It is currently infeasible and expensive toextract psilocybin from mushrooms, and even the best chemical synthesismethods require expensive and difficult-to-source starting substrates.

Accordingly, there is a need of methods to produce high efficiency,standardized preparations of the target compounds for medical use whileusing acceptable solvent systems to create a more consistent supplychain.

This background information is provided to reveal information believedby the applicant to be of possible relevance to the present invention.No admission is necessarily intended, nor should be construed, that anyof the preceding information constitutes prior art against the presentinvention.

SUMMARY OF INVENTION

The present invention is directed to an extraction process ofpsychoactive compounds from psychedelic fungus, for example, thePsilocybe cubensis species of psychedelic mushroom. The principalpsychoactive compounds in Psilocybe cubensis include psilocybin andpsilocin. In particular, the extraction process of psychoactivecompounds involves drying fresh Psilocybe cubensis, followed bygrinding, extraction with a solvent in one or more steps, one or moresteps of filtration, optional adjustment of the pH if the solvent isacidic (acid/water/alcohol) or alkaline (base/water/alcohol),evaporation of the solvent, and standardization. Optionally, the processincludes drying to result in a final powdered psilocybin mushroomextract.

This summary does not necessarily describe all features of theinvention.

Disclosed herein is a process for forming an extract of psychoactivealkaloids from psychedelic fungus comprising the steps of: soaking abiomass of dried, raw psychedelic fungus in a solvent selected from thegroup consisting of methanol and a water-methanol mixture in order todissolve the psychoactive alkaloids in the solvent; filtering anundissolved portion of the biomass from the solvent; evaporating thesolvent sufficiently to remove the methanol completely, leaving aconcentrated slurry or a residue that is converted to the concentratedslurry by adding water thereto: and standardizing the concentratedslurry by adding thereto a quantity of carrier measured to achieve aspecified purity of extract.

Also disclosed is a process for forming an extract with a specifiedconcentration of psychoactive alkaloids from a dried, raw, psychedelicfungus comprising the steps of: soaking a biomass of the dried, rawpsychedelic fungus in a solvent that is methanol, a water-methanolmixture or a buffered water-methanol mixture in order to dissolve thepsychoactive alkaloids in the solvent; filtering an undissolved portionof the biomass from the solvent; evaporating the solvent sufficiently toremove the methanol completely, leaving a concentrated slurry or aresidue that is converted to the concentrated slurry by adding waterthereto; measuring a psychoactive alkaloid content in the concentratedslurry; measuring a dry mass content in the concentrated slurry; usingthe psychoactive alkaloid content, the dry mass content and thespecified concentration to determine a quantity of a carrier to add tothe concentrated slurry in order to obtain the specified concentrationof the psychoactive alkaloids in the extract; standardizing theconcentrated slurry by adding thereto the quantity of the carrier; anddrying the concentrated slurry to result in the extract with thespecified concentration of the psychoactive alkaloids.

BRIEF DESCRIPTION OF DRAWINGS

The following drawings illustrate embodiments of the invention, whichshould not be construed as restricting the scope of the invention in anyway.

FIG. 1 is a high-level flowchart showing the key steps of a process forextracting psychoactive alkaloids from psilocybin fungus, according toan embodiment of the present invention.

FIG. 2 is a flowchart showing more detailed steps of a process forextracting psychoactive alkaloids from Psilocybe cubensis using a 75%ethanol solvent, according to an embodiment of the present invention.

FIG. 3 is a flowchart showing more detailed steps of a process forextracting psychoactive alkaloids from Psilocybe cubensis using ahydro-ethanol solvent, according to an embodiment of the presentinvention.

FIG. 4 is a flowchart showing more detailed steps of a process forextracting psychoactive alkaloids from Psilocybe cubensis using a watersolvent, according to an embodiment of the present invention.

FIG. 5 is a flowchart showing more detailed steps of a process forextracting psychoactive alkaloids from Psilocybe cyanescens using amethanol solvent, according to an embodiment of the present invention.

FIG. 6 is a flowchart showing more detailed steps of a process forextracting psychoactive alkaloids from Psilocybe cubensis using abuffered acidic solvent, according to an embodiment of the presentinvention.

FIG. 7 is a flowchart showing more detailed steps of a process forextracting psychoactive alkaloids Psilocybe cubensis using a bufferedalkaline solvent, according to an embodiment of the present invention.

FIG. 8 is a schematic diagram of the apparatus used for the extractionof psychoactive compounds according to an embodiment of the presentinvention.

DESCRIPTION A. Glossary

Psilocybin fungi, or psilocybin mushrooms—these are a group of fungithat contain at least one psychoactive alkaloid, and generally containpsilocybin and psilocin. They may also contain other psychoactivealkaloids such as baeocystin, norbaeocystin, ibotenic acid andnorpsilocin. The genera of these mushrooms include Copelandia,Gymnopilus, Inocybe, Panaeolus, Pholiotina, Pluteus, Amanita andPsilocybe.

Psilocybe mushrooms—these form a genus of gilled mushrooms in the familyHymenogastraceae. Most species contain the psychedelic alkaloidspsilocybin, psilocin and baeocystin.

Psilocybin—this is a psychedelic prodrug produced by numerous species ofmushrooms, collectively known as psilocybin mushrooms. Psilocybin isconverted by the body to psilocin, which has mind-altering effects suchas euphoria and hallucinations, but can also lead to nausea and panicattacks.

B. Process Overview

Referring to FIG. 1 , a flowchart is shown of the basic steps of theextraction process for extracting psychoactive compounds from psilocybinfungus. In step 100, a solvent is added to a biomass of dried andground, raw psilocybin fungus. The raw psilocybin fungus includesPsilocybe cubensis mushrooms, Psilocybe cyanescens mushrooms, Amanitamuscaria mushrooms or a mixture of these. Other species of psychedelicmushrooms may also be used.

The parts of the mushrooms used include, for example, caps, gills,stems, and hyphae, and more particularly, any part of the psilocybinmushroom or mycelium can be included. In other cases, the raw psilocybinfungus parts used include only caps, or only stems, or only gills, oronly hyphae or only mycelium or any mixture thereof. In still othercases, parts of the raw psilocybin fungus used are those that wouldnormally be considered waste, in which valuable psychoactive compoundsare found only in lower concentrations. The mushroom parts may be groundusing a milling machine or pulverization device, for example.

Ideally, the moisture content of the raw plant material after drying islow compared to the total dried biomass weight. For example, themoisture content may be under 5% for smaller scale extractions and under10% for larger scale extractions. Wet mushrooms, e.g. with a moistureabove 80%, will degrade rapidly. Dried biomass lends itself well toextraction since the drying process usually breaks down cell walls,allowing solvent to capture the molecules inside. The temperature of theoven and the drying time depend on how much moisture is in the rawpsilocybin fungus, and on the quantity of raw psilocybin fungus.

The solvent may be selected from a range of different solvents,including lower aliphatic alcohols (C=1, 2, 3 or 4), water,alcohol-water mixtures, strong alkaline buffers, and strong acidicbuffers. A wide range of solvent to solid ratios can be used. Typically,a 1 to 50:1 solvent-solid ratio (L:kg) may be used for the extraction.The amount of solvent used generally varies according to the weight ofthe raw psilocybin fungus.

In step 102, as a result of adding the solvent, and soaking the biomassof dried, raw psilocybin fungus in the solvent, essential elements orpsychoactive alkaloids found in the biomass dissolve into the solvent.The solvent may be at a low or high temperature, and pressure may beapplied to the solvent. In some embodiments the solvent is at roomtemperature. The optimal temperature of extraction varies depending onthe solvent type used for the process. However, the optimal temperaturefor extraction is in range of 5-95° C. The useful temperature rangespans most of the liquid state of the solvent used, and upper and lowerlimits are determined by physical practicalities and limits of theavailable apparatus. Still, the temperature of the solvent may beoutside of this range in other embodiments. The duration of theextraction is from 10 minutes to 12 hours, with or without agitation.Optimum duration is determined by experimentation, and depends on thechosen solvent and the strength of agitation in the extraction vessel.

If pressure is applied it may be in the range of 50 kPa-100 MPa aboveatmospheric (7-15000 psig). The lower limit of pressure is indicative ofwhen a benefit is seen in the rate at which the psychoactive alkaloidsdissolve in the solvent, since the increased pressure may increase thereaction kinetics of the dissolution of the psychoactive alkaloids intothe solvent. The upper limit is determined by what is physicallypractical given the constraints of equipment to safely operate underhigh pressure. Nevertheless, other pressures may be used. Solventcomposition, particle size and the temperature of extraction willdetermine how much pressure needs to be applied.

The extraction results in an extraction slurry, which is formed ofundissolved and insoluble solids from the mixture of biomass andsolvent, which now carries dissolved extract. Some of the undissolvedsolids may be undesirable components.

In step 104, the extraction slurry is filtered, resulting in a residue(i.e. the undissolved portion of the biomass) and filtrate. Thefiltering step may be carried out with the extraction slurry still hot,or it may first be allowed to cool. The extraction and filtration stepsmay be repeated multiple times on the same residue, with a fresh batchof solvent, which may have the same composition as the first solvent orit may be a different solvent.

In step 106, if the filtrate results from using a strongly acidic oralkaline solvent, then the filtrate is brought closer to neutral, e.g.to a pH between 4 and 9 or thereabouts. Desirable effects, such as morecomplete extraction, or preservation of the alkaloids fromdecomposition, or the ability to selectively extract certain specificalkaloids, are seen during the extraction stage when stronger acids oralkalis are used compared to weaker ones.

In step 110, evaporation of some or all of the solvent from the filtrateresults in a concentrated slurry 112 (liquid and solids) or just solids.If the solvent is methanol, then all of it is evaporated to reduce thelikelihood of toxicity. For other solvents, only some of the solventneeds to be evaporated. In the case where solids are obtained from theevaporation, water is added to the solids to form the concentratedslurry 112. The solids tend not to dissolve back into solution becausethey are less soluble in methanol and ethanol, for example, than water.Also, the solids may be less soluble in the colder water that is addedback than the warmer or hotter water that is used for the extraction.Another reason is saturation of the solution, or that some of the solidsare irreversibly precipitated.

In step 114, standardization of the concentrated slurry takes place. Theaim is to stabilize the extract by adding sufficient stabilizer (e.g.ascorbic acid and silica), and then titrating with a carrier such asmaltodextrin to result in a final, known concentration of psychoactivealkaloids. The slurry is analyzed for dry mass concentration andalkaloid content. The liquid component of the concentrated slurry isfirst analyzed using a loss-on-drying analysis and high performanceliquid chromatography coupled with diode array detection or massspectrometry to determine the alkaloid content. Depending on thedetermined alkaloid content, non-toxic carriers are added to theconcentrated slurry so as to provide a desired ratio between the weightof alkaloid and weight of carrier in the concentrated slurry. The addedcarriers, blending agents, excipients, flow aids etc. that may be usedinclude maltodextrin from corn, potato or tapioca for example, gumarabic, silicon dioxide, microcrystalline cellulose, ascorbic acid,sodium benzoate, sodium phosphate, sodium citrate, rice hulls, and rice.A combination of any of these carriers may be used.

In step 116, the concentrated slurry is dried to remove the remainingsolvent or water, resulting in a powdered psilocybin mushroom extractwith a known concentration by weight of psychoactive compound(s). Theextract is a powdered psilocybin mushroom extract that may have, forexample, a total psychoactive alkaloid concentration of 0.1-10% by dryweight. Other compounds may be included in the extract. These may besugars, proteins, carbohydrates and fats, and may make up about half ofthe extract. Step 116 is optional, as it may be the intention to producea liquid extract instead of a powdered extract.

C. Exemplary Embodiments 75% Ethanol Solvent

Referring to FIG. 2 , an exemplary detailed process is shown for theextraction of psychoactive compounds from Psilocybe cubensis mushroomsusing a 75% ethanol solvent.

In step 130, 2.5 kg of raw psilocybin mushrooms from the Psilocybecubensis species is provided. In step 132, the raw psilocybin mushroomsare dried in a forced air oven at 25° C., for 10 hours. The aim is todry the mushrooms so as not to significantly reduce their psychoactivealkaloid concentration. For example, if too high a temperature or toolong a time at a specific temperature were used, the alkaloids may startto decompose. The resulting, dried biomass is 140 g. In step 134, thedried biomass is ground using a hammer mill or the equivalent, to aparticle size of 200 mesh.

In step 136, a 5 kg quantity of the 75% (by weight) ethanol solvent,formed by mixing 3 parts of ethanol to 1 part of water by weight, isplaced in an extraction vessel. The dried, ground biomass is also placedin the extraction vessel, which is heat-controlled and agitated.

The extraction proceeds in step 140 as the biomass soaks in the solvent.The temperature of the extraction process is 70° C., and the duration ofextraction is 4 hours. The temperature remains constant during theextraction process.

In step 142, the resulting mixture of biomass solids and solvent withdissolved extract, is filtered while still hot, i.e. still at 70° C., orslightly lower due to ambient cooling. This removes a residue withundissolved psilocybin mushroom components from the filtrate. The filterused is a 10 μm sieve. The filtrate from this step is filtrate A. Instep 144, the residue is retained and placed back into the extractionvessel. In step 146, another 5 kg of 75% ethanol is added to theretained residue.

In step 150, the extraction process of the residue continues at the sametemperature as for the initial extraction step, i.e. at 70° C., for atime of 4 hours. Again, the temperature remains constant during theextraction process.

In step 152, the second resulting mixture, of biomass solids and solventwith dissolved extract, is filtered to remove the residue of unwantedsolid material. The filter used is a 10 μm sieve. Note that in otherembodiments a differently sized filter may be used here or in the priorfiltration step, or the liquid may be decanted from the residue withoutfiltering. In some embodiments, a centrifuge may be used to helpseparate the liquid from the residue. Filtrate B from the secondfiltration process may have a lower concentration of psychoactivecompounds than filtrate A from the first filtration step. Filtrates Aand B are then mixed in step 154 to result in bulk filtrate C. Moreextract can be obtained by splitting the solvent into two or morebatches and using each one sequentially to soak the biomass, compared tousing a single volume of solvent.

The bulk filtrate C is then processed with a rotary evaporator in step156 to remove solvent until the volume of filtrate C is 2.5 liters. Atthis point, the reduced amount of filtrate C is a concentrated slurry,due to the precipitation of water-insoluble components, for example.

The volume of 2.5 L is chosen because the mixture now has a low enoughethanol content that the carriers can be mixed in. By preferentiallyremoving ethanol over water, which occurs naturally during theevaporation, it also gives the later spray-drying step a lower risk ofexplosion compared to if a 75% ethanol slurry were sprayed directly.

In step 160, after some of the solvent has been removed using the rotaryevaporator, the concentrated slurry is then standardized. Thestandardization process uses a titration procedure to determine theconcentration of the psychoactive alkaloids in the concentrated slurry.The standardization procedure entails adjusting the concentration ofpsychoactive alkaloids the concentrated slurry to a desired target, suchas 1.00% by dry weight. In this example, 4.7 g of ascorbic acid, 1.9 gof SiO₂ and 47 g of maltodextrin are added to the concentrated slurry.

In step 162, after the standardization process, the standardizedconcentrated slurry is dried using a bench-top spray dryer. This resultsin 100 g of powdered psilocybin mushroom extract with a total alkaloidconcentration of 1.00% by weight. As can be seen, the purity of theextract can be defined as a percentage to a precision of two decimalplaces.

0-100% Ethanol Solvent

Referring to FIG. 3 , a process is shown for the extraction ofpsychoactive compounds from Psilocybe cubensis using a generalhydro-ethanol solvent. The solvent may range from a percentage of <1% ofethanol in water to 100% ethanol.

In step 180, 2.5 kg of raw psilocybin mushrooms from the Psilocybecubensis species is provided. In step 182, the raw Psilocybe cubensis isdried in a forced air oven at 25° C. for 10 hours. In step 184, theresulting dried biomass is ground in a hammer mill or the equivalent, toparticle size of 200 mesh.

In step 186, 5 kg of solvent, having a 0-100% ethanol concentration isadded to an extraction vessel into which the ground biomass is placed.The extraction vessel is an agitated, heat-controlled vessel.

In step 190, the extraction proceeds as the biomass is soaked. Thetemperature of the extraction is elevated above room temperature to 70°C. Temperature and pressure, if applied, are generally selected so thatthe solvent does not boil if elevated temperatures are used. Theduration of the extraction is 4 hours.

In the step 192, the extraction slurry is filtered to remove residuewith undissolved Psilocybe cubensis from the filtrate. The residue maybe treated with another extraction step if desired, and if so, thefiltrate from the subsequent step is combined with the filtrate from thefirst filtration.

In step 194, solvent from the filtrate is partially evaporated using arotary evaporator. The resulting concentrated slurry is then subjectedto a standardization process in step 196. The standardized concentratedslurry is then dried using a bench-top spray dryer in step 198 to resultin a powder with an accurately determined concentration by weight ofpsychoactive alkaloids.

100% Water Solvent

Referring to FIG. 4 , a detailed process is shown for the extraction ofpsychoactive compounds Psilocybe cubensis using 100% reverse osmosiswater as the solvent.

In step 210, 2.5 kg of raw psilocybin mushrooms from the Psilocybecubensis species is provided. In step 212, the raw Psilocybe cubensis isdried in a forced air oven at 25° C. for 10 hours. The dried biomass is140 g. Note that the dried biomass is the same weight in differentexamples because the mushrooms were from the same starting batch. Instep 214, the dried biomass is ground in a hammer mill or theequivalent, to a particle size of 200 mesh.

In step 216, 5 liters of solvent, which is 100% reverse osmosis water,is placed in an extraction vessel with the dried biomass, which isheat-controlled and agitated.

In step 220, the extraction proceeds. The temperature of the extractionprocess is 90° C., and the duration of the extraction is 12 hours. Inthe step 222, the extraction slurry is filtered while still hot toremove residue with undissolved Psilocybe cubensis from the filtrate.The filtrate from this step is considered as filtrate A. In step 224,the residue is retained and placed back in the extraction vessel. Instep 226, another 5 liters of 100% reverse osmosis water is added to theresidue. In step 230, the extraction process of the residue continues ata temperature of 90° C., for 10 hours. The temperature remains constantduring the extraction process. In step 232, the second resultingmixture, of biomass solids and water with dissolved extract, is filteredwhile still hot to remove the residue of unwanted solid material.Filtrates A and B are then mixed in step 234 to result in bulk filtrateC.

The bulk filtrate C is then processed with a rotary evaporator in step236 to remove solvent until the volume of filtrate C is 2.5 liters. Atthis point, the reduced amount of filtrate C is a concentrated slurry,due to the precipitation of some of the psychoactive alkaloids.

In step 240, after some of the solvent has been removed using the rotaryevaporator, the concentrated slurry is then standardized. Thestandardization process uses a titration procedure to determine theconcentration of the psychoactive alkaloids in the concentrated slurry.The standardization procedure entails adjusting the concentration of thepsychoactive alkaloids in the concentrated slurry to a desired drytarget. In this example, 6.3 g of ascorbic acid, 2.5 g of SiO₂ and 63 gof maltodextrin are added to the concentrated slurry.

In step 242, after the standardization process, the standardizedconcentrated slurry is dried using a bench-top spray dryer. This resultsin 140 g of powdered psilocybin mushroom extract with a total alkaloidconcentration of 0.50% by weight.

100% Methanol Solvent

Referring to FIG. 5 , a process is shown for the extraction ofpsychoactive compounds from Psilocybe cyanescens mushrooms using 100%methanol as the solvent.

In step 260, 2.5 kg of raw psilocybin mushrooms from the Psilocybecyanescens species is provided. In step 262, the raw Psilocybecyanescens is dried in a forced air oven at 25° C. for 10 hours. Thedried biomass is 140 g. In step 264, the dried biomass is ground in acutting mill or the equivalent, to particle size of 200 mesh. In step266, 5 kg of solvent, which is 100% methanol, is added to an extractionvessel, which is heat-controlled and agitated. The dries biomass is alsoadded to the extraction vessel.

In step 270, the extraction proceeds. The temperature of the extractionprocess is a constant 25° C., and the duration of the extraction is 4hours. A pressure of 100 kPa above atmospheric (15 psig) is applied tothe mixture of solvent and biomass during the extraction. In step 272,the extraction slurry is filtered to remove residue with undissolvedPsilocybe cyanescens from the filtrate.

The filtrate is then processed with a rotary evaporator in step 274 toevaporate all the methanol from the filtrate. In this embodiment, allthe solvent is removed at this stage because methanol is not regarded assafe for human consumption, and there should be no trace amounts of itremaining in the final product. In step 276, 1.25 liters of reverseosmosis water at room temperature is added to the solid that isremaining after the evaporation step, to form a concentrated slurry.

In step 280, the concentrated slurry is standardized. In this example,1.84 g of SiO₂ and 46 g of maltodextrin are added to the concentratedslurry. In step 282, the standardized concentrated slurry is dried usinga bench-top spray dryer. This results in 95 g of powdered psilocybinmushroom extract with a total alkaloid concentration of 1.50% by weight.

Acidic Solvent

Referring to FIG. 6 , a process is shown for the extraction ofpsychoactive compounds from Psilocybe cubensis mushrooms using abuffered acidic solvent. In step 300, 2.5 kg of raw psilocybin mushroomsfrom the Psilocybe cubensis species is provided. In step 302, the rawPsilocybe cubensis is dried in a forced air oven at 25° C. for 5-10hours. The dried biomass is 140 g. In step 304, the dried biomass isground in a hammer mill or the equivalent, to particle size of 200 mesh.

In step 306, 5 L of solvent is added with the dried biomass to anextraction vessel, which is heat-controlled and agitated. The solvent isa pH-adjusted, hydro-ethanol mixture. For its preparation, 44 g ofanhydrous citric acid is placed into a 5 L vessel with 1.25 L of reverseosmosis water followed by 3.75 L of ethanol. The contents are mixeduntil completely dissolved. The pH of this solution is between pH 1.8and pH 3.

In step 310, the extraction proceeds. The temperature of the extractionprocess is 30° C., and the duration of the extraction is 4 hours. Instep 312, the extraction slurry is filtered to remove residue withundissolved Psilocybe cubensis from the filtrate. The filtrate from thisstep is named filtrate A. In step 314, the residue is retained andplaced back in the extraction vessel. In step 316, another 5 L of thesame solvent is added to the residue. In step 320, the extractionprocess of the residue continues at a temperature of 30° C., for 4hours. The temperature remains constant during the extraction process.In step 322, the second extraction slurry is filtered, to remove theresidue of unwanted solid material. Filtrates A and B are then mixed instep 324 to result in bulk filtrate C.

Bulk filtrate C is then brought to a pH of 5 with 5M sodium hydroxide.The amount of the sodium hydroxide depends on the specific mushroommatrix extracted, and is not possible to predict accurately. ThepH-adjusted, concentrated slurry is then processed with a rotaryevaporator in step 330 to remove solvent until the volume of filtrate Cis 2.5 liters. At this point, the reduced amount of filtrate C is aconcentrated slurry, due to the precipitation of some of thepsychoactive alkaloids.

In step 332, the concentrated slurry is then standardized. In thisexample, 4.7 g of ascorbic acid, 1.9 g of SiO₂ and 47 g of maltodextrinare added to the concentrated slurry. In step 334, the standardizedconcentrated slurry is dried using a bench-top spray dryer. This resultsin 100 g of powdered psilocybin mushroom extract with a total alkaloidconcentration of 1.00% by weight.

Alkaline Solvent

Referring to FIG. 7 , a process is shown for the extraction ofpsychoactive compounds from Psilocybe cubensis mushrooms using abuffered alkaline solvent. In step 350, 2.5 kg of raw psilocybinmushrooms from the Psilocybe cubensis species is provided. In step 352,the raw Psilocybe cubensis is dried in a forced air oven at 25° C. for10 hours. The dried biomass is 140 g. In step 354, the dried biomass isground in a hammer mill or the equivalent, to a particle size of 200mesh.

In step 356, 5 liters of solvent is added with the biomass to anextraction vessel, which is heat-controlled and agitated. The solvent isa pH-adjusted, hydro-ethanol mixture. For its preparation, 200 g ofsodium hydroxide pellets are placed into a 5 L vessel, with 1.25 L ofreverse osmosis water followed by 3.75 L of ethanol. The contents aremixed until completely dissolved. The pH of this solution is between pH11 and pH 12.

In step 360, the extraction proceeds. The temperature of the extractionprocess is 30° C., and the duration of the extraction is 4 hours. Instep 362, the extraction slurry is filtered to remove residue withundissolved Psilocybe cubensis from the filtrate. The filtrate from thisstep is named filtrate A. In step 364, the residue is retained andplaced back in the extraction vessel. In step 366, another 5 L of thesame solvent is added to the residue. In step 370, the extractionprocess of the residue continues at a temperature of 30° C., for 4hours. The temperature remains constant during the extraction process.In step 372, the second extraction slurry is filtered, to remove theresidue of unwanted solid material. Filtrates A and B are then mixed instep 374 to result in bulk filtrate C.

Bulk filtrate C is then brought to a pH of 5 with sufficient 5Mphosphoric acid. The pH-adjusted concentrated slurry is then processedwith a rotary evaporator in step 380 to remove solvent until the volumeof filtrate C is 2.5 liters. At this point, the reduced amount offiltrate C is a concentrated slurry, due to the precipitation of some ofthe psychoactive alkaloids.

In step 382, the concentrated slurry is then standardized. In thisexample, 4.7 g of ascorbic acid, 1.9 g of SiO₂ and 47 g of maltodextrinare added to the concentrated slurry. In step 384, the standardizedconcentrated slurry is dried using a bench-top spray dryer. This resultsin 100 g of powdered psilocybin mushroom extract with a total alkaloidconcentration of 1.00% by weight.

D. Apparatus

Referring to FIG. 8 , an example of the apparatus is shownschematically. Raw psilocybin mushrooms are provided in a hopper 400,for example, and are released in batches into container 402. The rawfungal material is then dried in a forced air oven 404. The driedbiomass is placed into a grinder 406 for grinding.

After the drying and grinding steps, the ground biomass is placed in anagitated, heat-controlled extraction vessel 410. The vessel holds thebiomass and solvent 412, such as lower aliphatic alcohols, water,buffered acid or buffered alkaline, or a mixture thereof. The vessel maybe surrounded by an insulating wall 408. Alternately, there may be aninsulating jacket wrapped around the vessel. The insulating wall 408 orjacket helps to maintain the contents 412 under a constant temperature(T) between 5-95° C. The pressure (P) inside the extraction vessel 410may be regulated up to 100 MPa (15000 psig).

After the extraction, the bottom of the extraction vessel 410 is openedat outlet 414 and the extraction slurry is collected in container 420.The extraction slurry is then fed into filter 422. After filtration, thefirst filtrate leaves the filter 422 and is collected in container 424.The residue 430 is then fed back at R into agitated, heat-controlledvessel 410 and more solvent (S) is added. After the second extraction,the extraction slurry is collected in container 420 and is then fed intofilter 432 (or filter 422). After filtration, the second filtrate andsolvent mixture leaves the filter 432 and is collected in container 436.

After the two filtration stages, if there are two, the filtrates aremixed in container 440. Otherwise, if there is only a single filtrationstep, mixing is unnecessary. Neutralizer is added as necessary to thefiltrate in container 440. The extraction slurry, pH-adjusted wherenecessary, is then passed to rotary evaporator 442 in which all or partof the solvent is evaporated, depending on the embodiment. If all thesolvent is evaporated, then reverse osmosis water is added to the solidsremaining after the evaporation.

The concentrated slurry is then passed to container 444 and tested todetermine its alkaloid content, using a titration setup 446. Carriersare added to container 444 with the concentrated slurry, and mixed. Thestandardized slurry is then placed in a bench-top spray drier 450 toproduce psilocybin mushroom extract that is collected in container 452.

E. Variations

Other embodiments are also possible. While only specific neutralizingagents, food grade acids and food grade bases have been mentionedherein, other neutralizing agents, food grade acids and food grade basesmay be used.

In general, unless otherwise indicated, singular elements may be in theplural and vice versa with no loss of generality.

Temperatures that have been given to the nearest degree include alltemperatures within a range of ±0.5° C. of the given value. Likewise,numbers and percentages are specified to the nearest significant digit.Values of pH are specified to ±0.5.

While exemplary pH ranges are given in some examples, other pH rangesare possible.

Throughout the description, specific details have been set forth inorder to provide a more thorough understanding of the invention.However, the invention may be practiced without these particulars. Inother instances, well known elements have not been shown or described indetail and repetitions of steps and features have been omitted to avoidunnecessarily obscuring the invention. Accordingly, the specificationand drawings are to be regarded in an illustrative, rather than arestrictive, sense.

It will be clear to one having skill in the art that further variationsto the specific details disclosed herein can be made, resulting in otherembodiments that are within the scope of the invention disclosed. Stepsin the flowchart may be performed in a different order, other steps maybe added, or one or more may be removed without altering the mainoutcome of the process.

In other embodiments, other drying techniques, temperatures anddurations may be used. It is possible in other embodiments to grind thedried biomass to lower or higher particle size than 200 mesh. Forexample, grinding to a mesh size of 40 would work in some embodiments.The choice of solvent may have an impact on which mesh size to grind thedried mushrooms to. Note that, in other embodiments, the grinding step334 may take place before or after the drying step 332.

Water purified by other purification technologies may be used instead ofreverse osmosis water. In alternative embodiments the solvent is 0.02%to 1.5% acetic acid in water. In alternate embodiment, the solventcomprises 75% ethanol, 25% water and 0.1M sodium hydroxide. Inalternative embodiments the solvent is a hydro-methanol mixture, with amethanol content in the range of below 1% to 100%. The hydro-methanolbased extraction follows the same steps as the extraction with a mixtureof ethanol and water (FIG. 3 ), and may use lower soaking temperaturesdue to the lower boiling point of methanol. Also, the methanol/watermixture can be evaporated to dryness instead of the partial evaporationin step 194, for safety. If evaporated to dryness, the concentratedslurry is then formed by adding reverse osmosis water to the residualsolid. If not evaporated to dryness, the residual slurry is diluted, ifnecessary for ease of handling, by adding reverse osmosis water to formthe concentrated slurry. If not diluted, the residual slurry is used asthe concentrated slurry. The result of evaporating the methanol is aresidue that is either solid or a slurry. Furthermore, thehydro-methanol solvent may be buffered with a strong acid or a strongalkali, following the processes in FIGS. 6 and 7 . Again, however, thesolvent may be completely evaporated instead of partially (330, 380) inorder to fully remove the methanol, with reverse osmosis water beingadded to the solid to form the concentrated slurry. If the solvent isnot completely evaporated, it should be evaporated enough to remove allthe methanol and leave a residual slurry. The residual slurry mayoptionally then be diluted, for ease of handling, with reverse osmosiswater to form the concentrated slurry. If not diluted, the residualslurry is used as the concentrated slurry.

The solvent may also be propan-1-ol, propan-2-ol, a butanol isomer, or amixture of any or all of these with water, in any percentage ratio.

Any of the solvents described herein may be used with any of themushroom varieties that include psychoactive alkaloids, for exampleAmanita muscaria.

The process may be scaled up using larger quantities and modifiedapparatus.

The extraction process in other embodiments may use varying appliedpressures and temperatures, which vary during the soaking steps.

All parameters, dimensions, materials, quantities and configurationsdescribed herein are examples only and may be changed depending on thespecific embodiment. Accordingly, the scope of the invention is to beconstrued in accordance with the substance defined by the followingclaims.

1. A method for generating, from psychoactive organisms, a psychoactivealkaloid extract comprising psychoactive alkaloid, the methodcomprising: a. providing a biomass of the psychoactive organisms; b.contacting the biomass with 10 to 100 milliliters (mL) of solvent pergram (g) of the biomass, wherein the solvent has a pH of 1-4 and is anacid:methanol:water mixture; c. removing undissolved biomass from thesolvent to result in a psychoactive alkaloid liquid; and d. evaporatingthe solvent from the psychoactive alkaloid liquid to yield thepsychoactive alkaloid extract comprising the psychoactive alkaloid. 2.The method of claim 1, wherein a methanol concentration in the solventis between 25% and 75% by volume.
 3. The method of claim 1, wherein anacid concentration in the solvent is 5% by volume.
 4. The method ofclaim 1, wherein the acid is acetic acid.
 5. The method of claim 1,wherein the solvent has a pH of 2.5.
 6. The method of claim 1, whereinthe psychoactive alkaloid comprises phosphorylated alkaloid anddephosphorylated alkaloid.
 7. The method of claim 6 comprising, prior to(b): selecting desired yields of the phosphorylated alkaloid and thedephosphorylated alkaloid in the psychoactive alkaloid; and selecting amethanol concentration in the solvent between 25% and 75% by volume inorder to obtain the desired yields.
 8. The method of claim 7, whereinthe desired yields are between 15-80%.
 9. The method of claim 7, whereinthe methanol concentration is selected using an S-curve or reverseS-curve graph that relates the methanol concentration to the desiredyields.
 10. The method of claim 6, wherein the phosphorylated alkaloidis psilocybin and the dephosphorylated alkaloid is psilocin.
 11. Themethod of claim 1, wherein the psychoactive organisms are Psilocybemushrooms.
 12. The method of claim 1 comprising, following (c):contacting the undissolved biomass with a second solvent that has a pHof 1-4 and is another acid:methanol:water mixture; removing theundissolved biomass from the second solvent; and adding the secondsolvent to the psychoactive alkaloid liquid.
 13. The method of claim 1,wherein: the psychoactive organisms comprise phosphorylatablepsychoactive alkaloid and non-phosphorylatable psychoactive alkaloid;the psychoactive alkaloid in the extract comprises the phosphorylatablepsychoactive alkaloid and the non-phosphorylatable psychoactivealkaloid; and the phosphorylatable psychoactive alkaloid in the extractconsists of phosphorylated alkaloid and dephosphorylated alkaloid. 14.The method of claim 1, wherein the contacting is performed: at atemperature ranging from 5-95° C.; and for a time period ranging from10-720 minutes.
 15. The method of claim 1, wherein the contacting isperformed at a pressure ranging from 7 to 20,000 psi (50 kPa-138 MPa).16. The method of claim 1, wherein contacting the biomass is with 10 to50 mL of solvent per gram of the biomass.